Category: Histology

We can split the nervous system into 

1. the central nervous system, which includes the brain and spinal cord, and 
2. the peripheral nervous system, pretty much all the nerves that branch out from that. 

We’ll see some different structures and cell types depending on where we look, but the overall purpose of the nervous system is to send and receive electrical signals. Like the power lines that send electricity through a city, each nerve is made of clusters of smaller neuron cells. 

In a transverse cross section neurons look like little circles. 

And when we slice a nerve long ways for a longitudinal view and see the long axons running the length of the nerve. 

The naming for nerves sounds similar to muscle bundle naming. Remember how for muscles you have perimysium, epimysium and endomysium? Well in nerves, We have perineurium, epineurium and endoneurium. 

The cell body, or soma, has a nucleus inside. Branching out from there are any number of dendrites, branches that collect electrical impulses from other cells. They sum up at the axon hillock where an impulse will travel down the axon. The axon can be over 95% of the volume of the neuron cell — and they can be long like over a metre long. 

These axons are what we just cut open on the cross section and most of what we see on longitudinal sections. Finally, the neuron ends at the axon terminals. They send messages in the form of neurotransmitters to other cells through synapses. 

Some axons are thin, while some have a squishy layer around them that helps them transmit signals faster. It’s called a myelin sheath, so we say that those neurons are myelinated. And, we can find an endoneurium surrounding it. Depending on the layers, the sizes can differ too.

1. Myelinated type A – 4-20 micrometres – 70 to 120 metres a second
2. Type B – 1 to 4 micrometres
3. Type C – 0.2 to 1.5 micrometres – 0.5 to 2.5 metres per second

Not only can axons vary, but the branching pattern can vary too. 

1. Multipolar
   1. most common 
   2. brain, spinal cord
2. Bipolar
   1. Nose and retina
   2. they only send afferent, or sensory information
3. Unipolar
   1. Cell body and a single axon – dendrites

But neurons aren’t the only type of cell in the nervous system. We also have glial cells, essentially supportive cells. 

For instance, astrocytes support and protect our neurons by regulating the blood brain barrier, helping form synapses, and clearing excess neurotransmitters. They’re kind of hard to see with traditional light microscopes, so unless you have an electron microscope you probably won’t get quizzed on it. 

Oligodendrocytes are another fun one — they help make the myelin sheath around neurons in the brain and spinal cord, while Schwann cells make the myelin in the peripheral nerves. 

Quick summary, this all started with our bundles of neurons organised into peripheral nerves like electrical wires in a cable. But we still have some big deal nervous tissue to tackle — the central nervous system, including the brain and spinal cord. Luckily for us, we can get our bearings with the spinal cord similarly to how we did the peripheral nerves. 

The longitudinal section looks familiar, but different and the transverse cross section is super unique. 

This cross section shows two different colours to work with — which come from myelin status. Since those myelin sheaths are so fatty and fluffy, think of myelinated fibres like marshmallows that make up white matter, while those dense, slow, unmyelinated fibres are the grey matter. 

Since the grey matter falls into this shape, we label these segments horns. And we have anterior, lateral, and dorsal horns. 

But there’s another big component to the central nervous system, the brain. 

Let’s look at these two different colours, since their tissue level anatomy is different. Before we get to neurons, we have a few layers of connective tissue called the meninges. If you’ve heard of the disease meningitis, it’s inflammation of these layers. 

The most superficial layer is the dura mater, a layer of dense connective tissue that sticks to the skull. Deeper than that is the arachnoid layer which is thin and looks like spider webs, hence the name, and connects to the delicate thin pia mater underneath. 

And aside from some connective tissue around blood vessels, all the other structures of the brain can be classified as nervous tissue. But like I said, layers. The outermost layer of the cerebrum is the cerebral cortex, and deeper than that, the subcortical white matter. The cerebral cortex has 6 layers of its own and only a couple of cell types to differentiate between.

Most of our body mass is made of muscle and connective tissue like bone, fat, and liquids. 

Meanwhile, epithelial tissue doesn’t make up a ton of your body mass but still shows up in crucial spots around the body and does a handful of important jobs.

Epithelium’s two main jobs are

1. forming layers of cells that cover internal and external surfaces like the lining of your blood vessels and skin. 
2. secreting different substances either within the body or outside of the body. In some cases they’re responsible for forming the functional bulk of certain organs, what’s called parenchyma. For instance, the liver is 80% epithelial liver cell, or hepatocyte, by mass.

Epithelium can come in all kinds of different specialised tissue types that lets them do other unique jobs depending on where we find them, but most of the time we care about them as protective layers and secreting cells. 

So when we look at images of epithelial tissue, we have a big challenge: Epithelium is usually mixed in with all kinds of different tissue types, so our first job is to get our bearings and identify the tissue of interest. 

The cool thing about epithelial cells though is that they’re polar — they have distinct top and bottom poles that are oriented around a basement membrane which separates it from the structures around it. 

Between the cells and basement membrane is a layer of connective tissue that glues the cells to the membrane. It is called the lamina propria and it has blood vessels to feed the cells above. It touches the basal surface. 

The opposite surface is called the apical pole. Cilia and microvilli can be seen. (eg. lungs).

In the intestine the apical pole is towards a lumen. Lumen means the empty inner part of a tube. Lateral faces of the epithelial cells communicate through the gap junctions (ions), tight junctions (proteins), and desmosomes (anchor cells together).

Epithelial tissue is avascular. It gets its oxygen and nutrients by diffusion from the capillaries in the lamina propria. And lamina peoria supply stem cells to replace old epithelial cells. Our skin’s epithelium does this really quickly, which is why it grows back so fast if we scrape off a layer.

Naming epithelial Tissue

When naming, we consider the number of layers and cell shape. 

When we consider the shape we have

1. squamous cells which are squished flat like pancakes. Because of that, they have a squishy, oval shaped nucleus and not a whole lot of organelles within them. 
2. Cuboidal, or cubed shaped epithelium, which typically have large, round nuclei and plenty of organelles. 
3. columnar, or column shaped. A lot of those specialised epithelial cells like mucus secreting cells are columnar.

Once we know the shape of the cell, we need to arrange them on the basement membrane. That part of their organisation makes up the first part of their name.

1. If it’s organised into a single sheet, it’s called simple epithelium. 
   1. simple squamous epithelium which lines blood vessels. Its thinness makes it ideal for when you need to pass /some/ substances like oxygen through but still want it to keep blood contained. 
   2. simple cuboidal for secretion of substances like in the seminiferous tubules in the testis.
   3. simple columnar like in the walls of the gastrointestinal tract that can also secrete and absorb things. 
   4. The caveat is pseudostratified epithelium, which is technically a type of /simple/ columnar tissue, but it looks like it’s stacked into multiple layers.
2. stratified epithelium happens anytime you have two or more layers of epithelial cells. 
   1. You can have stratified simple, stratified cuboidal, and stratified columnar.
   2. In stratified epithelium, the farther away the cells get from that source of nutrients, the less nutrients they get. So we find a special type of stratified epithelium called keratinized epithelium, which is what it sounds like. Those epithelial cells are far away from the lamina propria, they die, they lose their nucleus and get filled with the tough protein keratin. If you’ve heard that your top layer of skin is all dead cells, it’s true, because it’s this stuff.
   3. transitional epithelium — stacks of different types of epithelial cells that transition from one cell type to another. We mostly see this in anatomy that needs to stretch — like bladder tissue, what sometimes gets called urothelium. 

Glandular Function of Epithelium

There are epithelial cells which, either pump things into the bloodstream or receive messages from the bloodstream.

We call these kinds of cells glandular epithelium, and when we get a bunch of them together, they can form full organs like the thyroid gland. We divide glands further based on how and where they release their message. 

1. They might send and receive messages outside the body, what are called exocrine glands, or 
2. within the body, what are called endocrine glands. If that name sounds familiar, it’s because the endocrine system is responsible for hormones.

When endocrine glands secrete hormones, they travel through the bloodstream, until they arrive at its target tissue, then they communicate their message. And these glands can show up in a lot of places. 

1. Like in the pineal gland in the brain. It’s home to nervous tissue like astrocytes, but also to epithelial cells called pinealocytes that pump out the hormone melatonin. 
2. You also have endocrine cells alongside other tissues. For instance, the cells responsible for making sperm are in the testes, but so are different cells called Leydig cells that secrete testosterone from the testes into the bloodstream.

But some glands don’t pump things into the body at all, exocrine glands pump them outside the body or into different cavities within the body. 

1. For instance, goblet cells in the GI tract secrete a protective layer of mucus out into the intestine, which is technically outside of our bodies. 
2. Or salivary glands — they secrete a bunch of enzymes and proteins into the saliva. Those enzymes aren’t communicating anything like a hormone would, so it’s an exocrine gland. 
3. Another example are apocrine glands, a subtype of exocrine gland that gives off odorants.  Something that carries a smell is more useful if it travels outside the body to a nose, so it’s an exocrine gland, not endocrine. 

One of the main places we see epithelial cells is in the digestive system. Therefore I will write more information in the digestive system post. These short notes are enough for this section.

The purpose of muscle is to produce force. Despite the type, the muscles produce force when they contract. But, they work slightly differently. And, their function is based on the form. 

Skeletal muscle

We can find in biceps, triceps, and squads, etc. They are optimised for quick and strong contractions. Because it helps our skeleton to move against gravity and heavy weights. 

When we see in slides, we have 2 views. 

1. Transverse cross section 
2. Longitudinal cross section ; parallel of the fibres, we see the sarcomere 

Whole muscle is covered by epimysium(dense connective tissue). Epi means upon. Mysium means muscle. In the muscle bulks, we see fascicles(bundles). When there is a bundle, we need something to hold it together. Each fascicle is held by perimysium. And Peri means around. Within these fascicles we can find muscle fibres coated with loose connective tissues called endomysium.  Endo means within. Inside the fasciculus we find many myofibres. Myofiber is covered by sarcolemma. Inside myofibers we can find myofibrils. They are held together by sarcoplasm. 

When we look at the muscles longitudinally, we find sarcomeres. The size is 2-3 µm. 

Cardiac Muscle

They are exclusive to the heart only. 

The heart is located between the lungs. The function is to pump blood. And the connective tissues help to separate the inside from the outside and keep it anchored to the place. 

Pericardium(parietal pericardium) covers the whole unit. It is a dense connective tissue. Inside this we have serous fluid. Then we have epicardium(visceral pericardium). Inside this we have myocardium. This is the muscle tissue we usually study in histology. Inner tissue layer is endocardium. It is a combination of epithelial tissues and connective tissues lining the chambers of the heart. 

Myocardium is the tissue. It is made of myocytes (cells) . 

Myocytes are rectangular in shape. One nucleus. Many mitochondria. As the heart has to contract simultaneously, there should be zero lag of electrical impulse. So, intercalated discs (gap junctions) allow the muscle cells to transmit signals quickly. 

Smooth Muscle

They surround organs that need to constrict and expand. Blood vessels, uterus, bladder, and gastrointestinal tract. 

They are called smooth because they don’t have striations. They are spindle shaped. They also have gap junctions. 

Cells are the smallest structural and functional unit of life. 

Tissue is a group of cells of similar function and origin that form functional units

An organ group of tissue adapted to perform specific functions

An organ system is group of organs work together to perform more functions

Organism

Tissue preparation for light microscope

Tissue sampling

• Histological examination of tissues starts with surgery, biopsy or autopsy (or necropsy).
• When collecting the samples clinical details and adequate specimens are important

Fixation

• To preserve
• Fixatives
• 10% formaldehyde
• 1mm/hour
• Wax processing

Dehydration and clearing

• Graded solutions of alcohol 50 – 100%
• 20 – 30 minutes each

Clearing

• Dealcoholation
• Xylene and chloroform

Embedding

• Paraffin wax
• Automatic tissue processor

Sectioning

• Microtome
• 3 – 10 micrometers
• Water Bath
• Frozen specimens – liq. Nitrogen or rapid freeze bar – pre cooled steel blade and glass 

Mounting and staining the sections

• Basophilic
• Acidophilic
• Hematoxylin and Eosin – reverse order process – nuclear dark purple – cytoplasm and intracellular structures  – pink 
• Hematoxylin – basophilic – dark blue
• Eosin – acidophilic – pink 

Covering

• Coverslip – permanently affixed – mounting medium

Light Microscope

Parts ; 

1. Eyepiece lens
2. Tube
3. Arm
4. Base
5. Illuminator
6. Stage
7. Revolving nosepiece
8. Objective lenses
9. Condenser lens
10. Coarse & fine focus
11. Diaphragm

Histochemistry & Cytochemistry

Methods for localizing cellular structures in tissue sections using unique enzymatic activity present in those structures. 

Perl prussian blue reaction – iron deposits in hemochromatosis 

Immunohistochemistry 

Histological + immunological + biochemical techniques for identification of specific tissue components (antigen/antibody).

Frozen sections are commonly used. In some cases paraffin wax.

Assays – cells on slides – or tissues (frozen / paraffin)

Frozen

• Unfixed 
  • Advantage ; antigens are unaltered
  • Disadvantage ; sections fall out during staining
• Acetone fixed
  • Precipitate proteins on to cell surface
  • CD antibodies
• Paraformaldehyde fixed
  • Freshly made / frozen asap

Paraffin Embedded

• Deparaffinized
• Rehydrated (graded alcohol 100% – 50%, then PBS)
• Antigen retrieval 
  • Treat with proteases ; to expose buried antigenic epitopes 
  • Heat in
    • Low pH citrate buffer
    • High pH EDTA buffer

Antigen Detection

1. Raising Antibodies
   1. Polyclonal antibodies ; multiple epitopes and several antibody types
   2. Monoclonal antibodies ; single epitope and produced by a single clone
2. Labeling Antibodies
   1. Fluorochromes 
   2. Cronochromes enzymes
   3. Electron Scattering Compounds

Method

1. Direct ; Tissue antigen + Labeled antibody
2. Indirect ; Tissue antigen + Primary antibody + Secondary antibody (LBL)
3. PAP ; Peroxidase Anti-Peroxidase method 

Applications

• Cancer Diagnostics
• Differential Diagnosis
• Treatment of cancer
• Research

General Immunochemistry Protocol 

1. Tissue Preparation
   1. Fixation
   2. Sectioning
   3. Mount Preparation
2. Pre-treatment
   1. Antigen Retrieval 
   2. Inhibition of indigenous tissue components
   3. Blocking of nonspecific sites
3. Staining
   1. Specimen, primary antibody, degree of sensitivity, processing time

Types of tissues

When we look into the microscopic version of the human anatomy we find many colorful stained pictures that look similar. But, they are not. There are 4 main tissue types that make up every other tissue of the body. 

1. Muscle 
2. Nervous
3. Epithelial 
4. Connective

Muscle Tissue

Skeletal Muscle ; Long. parallel fibers or striations. Multiple nuclei per fiber. Dark dots.  

Cardiac Muscle ; Exclusive to the heart. Rectangular in shape. Nucleus per cell. We can see intercalated disks. They are passages from cell to cell. As they have to send signals quickly. Fast contraction.

Smooth Muscle ; Blood vessels, Uterus, and Bladder. Contract simultaneously. Smallest muscle cells and jumbled up as sheets. 

Nervous Tissue

Brain, Spinal cord and the peripheral nerves. We have; 

• neurons (Main cells, Transmit nervous impulse in brain or along nerve) 
• Glial cells (Astrocytes, Schwann cells, satellite cells)

Epithelial Tissue

Skin and Borders between different organs. They are named based on the

• Layer ; smooth, stratified
• Shape ; Cuboidal, squamous, columnar, 
• Transitional. Around bladder and urethra (organs stretch)

Connective Tissue

Everything else is connective tissue. We have tendons, ligaments, fat, blood, cartilage, and bone.

3 types

• Loose ; less ground substance
• Dense ; more ground substances, thick fibres
• Specialized ; chondrocytes(collagen cells), osteoblasts(build bone), osteoclasts (resorb bone), adipocytes (store energy as fat)